Plasma membrane (the outer membrane) of intact eukaryotic cells is a phospholipid bilayer, characterized by an asymmetrical distribution of its phospholipid constituents between the inner and outer membrane leaflets. Thus, the aminophospholipids phosphatidylserine (PS) and phosphatidylethanolamine (PE), (particularly PS) reside almost entirely in the inner leaflet of the plasma membrane. This asymmetry of phospholipid distribution is important for various cellular functions, including normal cell interaction with its environment. This phospholipid asymmetry of plasma membrane is normally maintained, in an energy-dependent manner, by at least two systems comprising (1) an inward-directed pump, called the aminophospholipid translocase, which specifically and continuously moves PS and PE from the outer to the inner membrane leaflet in an ATP-dependent manner, while being inhibited by Ca2+, and (2) an outward directed floppase, that moves mainly PS from the inner to the outer membrane leaflet.
While maintenance of this membrane organization is fundamental to normal cell physiology, cells undergoing apoptosis or other modes of cell death, and cells undergoing activation (e.g., platelets during the process of blood clotting) manifest a dramatic process of “scrambling” of membrane phospholipids, with loss of their asymmetrical distribution and exposure of PS on the cell surface. The major factors responsible for this phenomenon comprise inhibition of the aminophospholipid translocase, and activation of phospholipid scramblase, a powerful, Ca2+ dependent protein system facilitating rapid bi-directional movement of phospholipids between the membrane leaflets.
Scrambling of membrane phospholipids and exposure of PS on the cell surface plays an important role in recruitment of macrophages for engulfment and removal of the apoptotic cells. In addition, phospholipid scrambling occurring in platelets plays an important role in rendering their surface a powerful catalytic platform for the assembly of several clotting factor complexes, such as the tenase and prothrombinase complexes. Accordingly, a strong correlation has been recently drawn between phospholipid scrambling and potent procoagulant effects exerted by apoptotic cells.
While these alterations have an important role in normal physiology, they may also be a factor in the pathogenesis of medical disorders associated with an abnormal, excessive blood clotting in numerous disorders such as arterial or venous thrombosis or thrombo-embolism (e.g., cerebral stroke, myocardial infarction, deep vein thrombosis, etc).
In addition to the plasma membrane of the cell, which manifests exposure of PS and an increased level of PE on the cell surface (i.e. on the outer leaflet of the plasma membrane) only upon phospholipid scrambling, many intracellular membranes (e.g., membranes of the endoplasmic reticulum) manifest such features on their surface as part of their normal structure. Cells wherein the plasma membrane is a PS-exposing membrane may be, for example, apoptotic cells or activated platelets. By contrast the plasma membrane of normal, healthy cells is not a PSE membrane, and thus these cells are not considered PSE cells.
It is desirable to have pharmaceutical agents, which have low molecular weight, and are capable of selective targeting, binding and potential accumulation within PSE cells.